Propofol composition containing sulfite

ABSTRACT

Sterile pharmaceutical compositions for parenteral administration containing 2,6-diisopropylphenol (propofol) are described for use as anesthetics. The compositions comprise an oil-in-water emulsion of propofol additionally comprising an amount of sulfite sufficient to prevent significant growth of microorganisms for at least 24 h after adventitious contamination.

This application is a continuation of application Ser. No. 09/247,449,filed Feb. 10, 1999, now U.S. Pat. No. 6,147,122, which is acontinuation-in-part of application Ser. No. 09/021,671, filed Feb. 10,1998 now abandoned, all of which are hereby incorporated by reference intheir entirety.

FIELD OF THE INVENTION

In one aspect, the present invention relates to new pharmaceuticalcompositions containing 2,6-diisopropylphenol, known as propofol, andsulfite. In another aspect, the present invention relates to the use ofthese compositions to induce anesthesia in mammals, including sedation,and the induction and maintenance of general anesthesia. In yet anotheraspect, the present invention relates to the use of sulfite as apreservative for parenterally administered oil-in-water emulsions, ingeneral. In still another aspect, the present invention relates to aprocess for the manufacture of oil-in-water emulsions containing sulfiteas a preservative.

BACKGROUND OF THE INVENTION

2,6-Diisopropylphenol, generically named propofol, is a well-known andwidely-used, injectable anesthetic with hypnotic properties used both asa sedative, and to induce and maintain general anesthesia. It is sold asDiprivan (trademark Zeneca) for human use and Rapinovet (trademarkZeneca) for veterinary use. Propofol is administered directly into thebloodstream either by bolus injection or by infusion. Because the onsetof anesthesia is largely controlled by a drug's diffusion rate throughthe blood-brain barrier, propofol's lipophilicity is key to its rapidactivity. This lipophilicity, however, renders propofol relativelyinsoluble in water, hence it must be administered in conjunction withsolubilizing agents, surfactants, or solvents; or as oil-in-wateremulsions (Jones et al. (1998) U.S. Pat. No. 5,714,520). All referencescited herein are incorporated by reference in their entirety.

As a parenterally administered agent, sterility of propofol formulationsis essential. Commercial formulations are oil-in-water emulsionscontaining approximately 1%-2% propofol in 10% soybean oil. Theseformulations also typically contain a surfactant, 1.2% egg phosphatidefor example, 2.25% glycerol to make the formulation isotonic, sodiumhydroxide to adjust the pH to physiological pH, and 0.005% EDTAequivalent (as edetate) to retard microbiological growth (all weightsapproximate) (Id.). Edetate containing formulations are notantimicrobially preserved by USP standards; however, microbial growth isretarded (Id.).

Non-preserved, propofol oil-in-water emulsion formulations havesignificant drawbacks arising from the fact that these formulationssupport microbial growth: strict aseptic handling technique is required;maximum utility time is 12 h maximum after vial entry. Handlingrecommendations include immediate administration after vial entry, anddisposal of infusion assemblies and of unused material after 12 h.Nevertheless, reports of nosocomial infections resulting fromadventitious contamination are not uncommon (Bennett et al. (1995) N.Engl. J. Med. 333:147-154). Improper handling techniques include delayedadministration after transfer from vial to syringe, use of 50- and100-mL products as multi-use, for multiple patients, and use of 50 and100 mL products for an extended time period.

An application for which preserved propofol formulations areparticularly advantageous is their use as a long-term sedative bycontinuous infusion. The risk of microbial contamination ofnon-preserved propofol in infusion devices increases both withresidency-time in the infusion device, and with increased manipulationof the device. The utility time of formulations containing EDTA salts(edetates) is at least 24 h compared to 6 to 12 h for non-preservedformulations (Jones et al.). A longer lasting formulation means thatfewer manipulations are required. The consequent reduced manipulationaccrues a number of important benefits: reduced probability of microbialcontamination, reduced probability of operator error, reduced drugwaste, and reduced labor intensiveness—all of which combine to increasesafety and reduce costs.

SUMMARY OF THE INVENTION

An extensive and vigorous evaluation of known antimicrobial agents forparenteral products led to the unexpected discovery that sulfite can beincluded in an oil-in-water emulsion of propofol in a non-toxic amountwhich is soluble in the aqueous phase and does not partition into theorganic phase, and which retards or suppresses the of growth of likelymicrobial contaminants, without destabilizing the emulsion and withoutadversely reacting with other formulation components. These results areespecially surprising in light of published data indicating that sodiummetabisulfite is completely ineffectual for this particular application(1% Diprivan (Zeneca) 0.1% Na₂S₂O₅) (Jones et al.)

Sodium metabisulfite is a salt of a sulfurous acid (formally,metasulfurous acid). The present invention includes all pharmaceuticallyacceptable derivatives of sulfurous acid (orthosulfurous acid) andmetasulfurous acid approved by the FDA for human use (sulfites) and anycombinations thereof. These compounds include, but are not limited to,sodium sulfite, sodium bisulfite, potassium sulfite, potassiumbisulfite, sodium metabisulfite, and potassium metabisulfite.

Accordingly, the present invention provides a sterile composition forparenteral administration comprising an oil-in-water emulsion in whichpropofol is dissolved in a water-immiscible solvent that is emulsifiedwith water wherein said emulsion is stabilized by means of a surfactant.The composition further comprises an amount of a sulfite sufficient toexhibit antimicrobial activity against microorganisms most likely tocontaminate the propofol preparation.

The present invention also includes the use of sulfites as preservativesfor any sterile, parenterally administered oil-in-water emulsion. Inaddition to propofol compositions, such formulations includetotal-parenteral-nutrition formulations, or oil-in-water vehicles forother pharmaceutical or therapeutic agents.

Additionally, the present invention includes a process for themanufacture of sterile, propofol oil-in-water emulsions for parenteraladministration comprising propofol dissolved into a water-immiscibleliquid emulsified with water, wherein said emulsion is stabilized bymeans of a surfactant and further comprising effective amounts ofsulfite as a preservative. Timing of the addition of the sulfite andcontrol of the process temperature are both critical to the maintenanceof antimicrobial activity in the composition. This aspect of theinvention may be advantageously applied to other drugs formulated as anoil-in water emulsion.

DEFINITIONS

In accordance with the present invention and as used herein, thefollowing terms are defined to have the following meanings, unlessexplicitly stated otherwise:

The term “edetate” refers to an anion derived from deprotonation ofEDTA. EDTA is a tetrabasic acid, thus an edetate may be mono-, di-, tri-or tetraanionic. The term “edetate” may also refer to a salt of anedetate anion.

The term “oil-in-water emulsion” refers to a distinct two phase systemthat is in equilibrium and in effect, as a whole, is kinetically stableand thermodynamically unstable.

The term “preservative” refers to an agent or agents that suppress orprevent microbiological growth at 24 h by no more than 10-fold comparedto time-zero.

The term “sulfite” refers to all pharmaceutically acceptable derivativesof sulfurous acid (orthosulfurous acid) and metasulfurous acid approvedby the FDA now or in the future for human use. These compounds includesodium sulfite, sodium bisulfite, potassium sulfite, potassiumbisulfite, sodium metabisulfite, and potassium metabisulfite.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1. Flow chart for the manufacturing process for sterile propofoloil-in-water emulsion formulations containing sodium metabisulfite.

DETAILED DESCRIPTION OF THE INVENTION

Development of a preservative-containing, oil-in-water emulsionformulations is a daunting task. Key requirements for the preservativeinclude:

a. Soluble in the aqueous phase, does not partition into the organicphase;

b. Low toxicity, since large volumes are commonly used;

c. Retardation/suppression of growth of likely microbial contaminants;

d. Compatibility with all other formulation components;

e. Not destabilizing of the emulsion.

Many currently used preservatives are lipophilic, and hence, would beineffective for use in oil-in-water emulsions because of requirement a.Emulsion physical stability and clinical performance depend criticallyon the particle-size distribution and the number of large particles(requirement e) (Dabbah et al. (1995) in USP OpenConference-Microbiological Compendial Issues The United StatesPharmacopeial Convention, pp 87-96).

Sodium metabisulfite, sodium bisulfite, sodium sulfite, potassiummetabisulfite, and potassium sulfite are used in many parenteralformulations as antioxidants and/or antimicrobials in concentrations of0.025-0.66%. The antimicrobial activity, however, requires a pH in the2.5-5.5 range. Oil-in-water emulsions, on the other hand, are typicallyformulated at pH 6-9 to assure the ionization of the headgroups of thephospholipid surfactants incorporated therein. The resultingelectrostatic repulsion favors the formation of small oil particles anddiscourages their coalescence with time. We have discovered stableemulsions containing each of the above sulfite-containing compounds inthe 4.5-6.4 pH range that nevertheless exhibit antimicrobial activity.We have also discovered a process for the manufacture of these emulsionswhich minimizes the loss of the sulfite-containing compounds throughautooxidation.

While not wishing to be limited to this mechanism, the inventors believethat the inhibition and destruction of microorganisms by sodiummetabisulfite, sodium bisulfite, sodium sulfite, potassiummetabisulfite, and potassium sulfite is highest at lower pH values. Theprimary activity of these compounds is believed to be due to theformation of sulfurous acid (H₂SO₃) and bisulfite ions. Theantimicrobial action is believed to be due to inhibition of enzymesystems, especially through oxidation of SH groups in enzymes andproteins. The bisulfite ions may also interact with pyrimidine bases(Foegeding, P. M. and Busta, F. F. “Chemical Food Preservatives,” pp802-832, in Disinfection, Sterilization, and Preservation, fourthedition, Ed S. S. Block, 1991, Pub. Lea and Febiger, William andWilkins, Philadelphia, USA).

1. Pharmaceutical Compositions

The composition of the present invention comprises a sulfite, preferablya salt of metasulfurous acid. More preferably, the sulfite is sodiummetabisulfite and other sulfurous acid salts such as sodium bisulfite,sodium sulfite, potassium metabisulfite, or potassium sulfite. Thesulfite will typically be present from about 0.0075% to about 0.66%weight. Preferably, the sulfite is present in the range of about 0.0075%to about 0.1% weight and most preferably about 0.025% weight. It will beapparent to one skilled in the pharmaceutical arts that other sulfitesmay be used in these compositions and that their weight percentages willdepend on the particular sulfite used.

Maximal dosages of sulfite will occur in long-term infusion situations,for example when used as a sedative. Typical dosages of propofol are0.3-3 mg/kg/h, but may range to 10 mg/kg/h in exceptional cases,equivalent to 1.68 L emulsion/day/70 kg. Under these conditions, thetotal sulfite administered is well below the limit set by the WorldHealth Organization (WHO) (7.0 mg/kg as SO₂) and is below the amountinfused in total-parenteral-nutrition amino acid formulations, as wellas during peritoneal dialysis (Gunnison and Jacobsen (1987) Crit. Rev.Toxicol. 17:185-214).

The composition of the present invention typically comprises 0.1 to 5%weight propofol. Preferable compositions comprise from about 1% to about2% weight propofol. More preferable compositions are about 1% weight andabout 2% weight propofol. The propofol may be dissolved in apharmaceutically acceptable water-immiscible solvent and emulsified inwater and said emulsion stabilized by means of a surfactant; or thepropofol may itself be emulsified in water without addition of awater-immiscible solvent and said emulsion stabilized by means of asurfactant.

Water-immiscible solvents suitable for the preparation of oil-in-wateremulsions suitable for parenteral administration are known to thoseskilled in the pharmaceutical arts (Handbook of Pharmaceutical ExipientsWade and Weller, Eds. (1994) American Pharmaceutical Association, ThePharmaceutical Press: London, pp 451-453). Typically, thewater-immiscible solvent will be a vegetable oil: for example, soybean,safflower, cottonseed, corn, sunflower, arachis, and castor. Thewater-immiscible solvent may also be a wholly or partially manufacturedmaterial, for example mono-, di-, and triglycerides, fatty acid esters,or chemically and/or physically modified vegetable oils. The presentinvention may also comprise any combination of said water-immisciblesolvents. When used, the water-insoluble solvent comprises up to about30% weight of the composition, preferably in the range of about 5% toabout 25% weight, more preferably in the range of about 10% to about 20%weight, most preferably about 10% weight.

The composition of the present invention comprises a pharmaceuticallyacceptable surfactant which aids in the emulsification of thewater-immiscible phase in water and stabilizes said emulsion (Id.).Suitable surfactants include naturally occurring surfactants: forexample, egg or soy phosphatides, either in their native or modifiedforms; manufactured non-ionic surfactants, for example a polyethyleneglycol or esters thereof; or any mixture thereof. Preferable surfactantsare egg or soy phosphatides, for example egg-yolk phospholipid. Theamount of surfactant effective in producing and maintaining a stableoil-in-water emulsion will depend the particular formulation. Thefactors and their relationships are well known to skilled practitionersin the pharmaceutical arts. These factors include the presence orabsence of a water-immiscible solvent, the particular water-immisciblesolvent used, the particular surfactant employed, the presence of salts,and the pH of the composition.

Preferably, the total number of fat globules/0.1 mL in the 1-20 μmparticle size range is ≦600,000 and in the 5-20 μm particle size rangeis ≦200,000. Preferably, the % volume of total fat globules/0.1 mL inthe 1-20 μm particle size range is ≦0.3 and is ≦0.2 in the 5-20 μm sizerange. Preferably, the mean fat globule size is less than 500 nm, andmore preferably less than 250 nm.

The composition of the present invention is formulated with pH in therange of about 4.5 to about 6.4. The pH may be adjusted as required bymeans of addition of an alkali, for example sodium hydroxide, or anacid, for example hydrochloric acid.

The composition of the present invention may be made isotonic with bloodby incorporation of a suitable tonicity modifier, for example glycerin(Id.).

The compositions of the present invention are sterile, aqueousformulations and are prepared by standard manufacturing techniquesusing, for example, aseptic manufacturing methods and sterilization byautoclaving.

Compositions of the current invention may be formulated to matchcommercial formulations in clinical performance and physical properties.Tables 1 below compares the composition of the preferred embodiment ofthe present invention with Diprivan. Table 2 below compares the physicalproperties of these two formulations.

TABLE 1 Comparison the formulation of a composition of the presentinvention with a commercial formulation. Propofol DIPRIVAN InjectableInjectable Component Emulsion 1% Emulsion 1% Propofol, mg/mL 10 10Soybean oil, mg/mL 100 100 Glycerin, mg/mL 22.5 22.5 Egg-yolkphospholipid, mg/mL 12 12 Disodium edetate, mg/mL — 0.05 Sodiummetabisulfite, mg/mL 0.25 — WFI q.s. to 1 mL pH 4.5-6.4 7.0-8.5 Diprivantrademark of Zeneca.

TABLE 2 Comparison of physical properties of a composition of thepresent invention with a commercial formulation. Propofol DIPRIVANPhysico-Chemical Injectable Injectable Parameter Emulsion 1% Emulsion 1%Appearance White emulsion White emulsion with no visible with no visibleoil droplets oil droplets Density 0.995 0.995 Osmolality, mg/mL 300 300Viscosity, centistokes 1.6 1.6-1.7 Diprivan trademark of Zeneca.

The following Table 3 shows that the particle size distribution of theemulsion containing sodium metabisulfite, sodium bisulfite, sodiumsulfite, postassium metabisulfite, or potassium sulfite are comparableto the Zeneca product containing 0.005% EDTA. The data show that theaddition of any of the salts did not cause an increase in the number oflarge particles which is a concern for injectable emulsions.

The size of particles with a submicron diameter (<1 μm) is monitoredusing Nicomp 370, manufactured by Particle Sizing Systems, SantaBarbara, Calif. This instrument measures an apparent average particlesize and distribution. The size and the number of fat globules largerthan lpm are determined using the AccuSizer™ 770 manufactured byParticle Sizing Systems, Santa Barbara, Calif. This technique allowsseparate monitoring of the number of particles and % volume of the oiltaken by these particles for the 1-20 μm and 5-20 μm diameter ranges.

TABLE 3 Total Number of Fat % Volume of Total Fat Globules/0.1 mLGlobules/0.1 mL Mean Fat Particle Particle Particle Size Particle SizeGlobule Size Size 1-20 μm Size 5-20 μm 1-20 μm 5-20 μm nm Diprivan312,911 ± 17,095 ± 6,398 0.050 ± 0.019 0.025 ± 0.010 194 ± 6 Injectable117,479 Emulsion 1% (Average of 3 different lots) Propofol 311,894 ±3,122 ± 642 0.022 ± 0.005 0.009 ± 0.002 198 ± 2 Injectable 71,940Emulsion 0.025% Sodium Metabisulfite (Average of 2 different lots)Propofol Injectable Emulsion (0.025%) of: Sodium 100,691 3,347 0.0120.004 201 Bisulfite Sodium Sulfite 128,338 6,689 0.021 0.010 239Potassium 156,569 6,470 0.022 0.011 197 Metabisulfite Potassium 50,1511,716 0.006 0.005 202 Sulfite

The compositions of the present invention are useful as anestheticsincluding sedation, and induction and maintenance of general anesthesia.Thus, in another aspect, the present invention provides a method forinducing anesthesia in mammals which comprises parenteral administrationof a sterile, aqueous pharmaceutical composition comprising anoil-in-water emulsion in which propofol, either alone or dissolved in awater-immiscible solvent, is emulsified in water, wherein said emulsionis stabilized by means of a surfactant; which further comprises aneffective amount of sulfite.

Dosage levels appropriate for the induction of desired degree ofanesthesia, for example sedation, or induction of or maintenance ofgeneral anesthesia, by the compositions of the present invention willdepend on the type of mammal under treatment and the physicalcharacteristics of the specific mammal under consideration. Thesefactors and their relationship in determining this amount are well knownto skilled practitioners in the medical arts. Approximate dosage levelsmay be derived from the substantial literature on propofol, may betailored to achieve optimal efficiency, and will be contingent on myriadfactors recognized by those skilled in the medical arts includingweight, diet, and concurrent medication.

The antimicrobial effects of sulfites may also be advantageously appliedto other sterile, oil-in-water emulsions for parenteral administration.Examples include total-parenteral-nutrition formulations andoil-in-water emulsions of other pharmaceuticals or therapeutic agents.

Oil-in-water emulsion total-parenteral-nutrition formulations areadministered by infusion to patients for whom oral nutrition isimpossible, undesirable, or insufficient. The emulsified lipids providea concentrated caloric content. These formulations may also containother nutrients, for example amino acids, vitamins, and minerals.Commercial examples of such formulations include Intralipid (trademarkPharmacia), Lipofundin (trademark Braun), and Travamulsion (trademarkBaxter). Accordingly, the present invention provides a steriletotal-parenteral-nutrition formulation comprising lipids or fatsemulsified in water which further comprises an effective amount ofsulfite as a preservative.

A wide variety of current and potential pharmaceutical or therapeuticagents are highly lipophilic, for example steroids, prostaglandins,leukotrienes, and fat-soluble vitamins. Such compounds may beadvantageously administered in oil-in-water emulsion vehicles comprisinga sulfite as a preservative, particularly when administration will occurover an extended period. Accordingly, the present invention provides asterile, therapeutic composition comprising a lipophilic pharmaceuticalor therapeutic agent, either alone or dissolved in a water-immisciblesolvent, emulsified in water, which further comprises an amount ofsulfite effective as a preservative.

2. Process for Manufacture

A scheme for the manufacture of compositions of the present invention isshown in FIG. 1. The present invention provides a process formanufacturing the compositions of the present invention comprising thesteps of:

1. Preparing an aqueous phase by adding glycerin and sodium hydroxideinto about 80% WFI in a compounding tank while maintaining thetemperature at approximately 40° C.;

2. Adding the egg-yolk phospholipid to said aqueous phase;

3. Homogenizing said aqueous phase;

4. Filtering said aqueous phase through a 5.0 μm filter;

5. Preparing an oil phase by dissolving propofol in soybean oil;

6. Filtering said oil phase through a 0.45 μm filter;

7. Combining and homogenizing said aqueous and oil phases;

8. Adding a solution of a sulfite compound dissolved in WFI near the endof the homogenization step;

9. Adding sodium hydroxide or hydrochloric acid solution to adjust thepH;

10. Adjusting to specified volume with WFI;

11. Microfluidizing the crude emulsion to the target globule size andparticle size distribution while maintaining the temperature at about30° C.;

12. Filtering the propofol oil-in-water emulsion into a filling vessel;

13. Filling and sealing containers under nitrogen;

14. Autoclaving said containers.

Typically, 12 mg/L of sodium hydroxide is added in step 3. Preferably,all steps are performed under a nitrogen atmosphere.

Timing of the addition of the sulfite is critical. If dissolved in theaqueous phase in step 3 or 4, the antimicrobial activity is lost,presumably from loss of bisulfite during processing. Optimalantimicrobial activity was obtained when the sulfite was added at step10. Typically, the sulfite is added by a stock solution of about 54 g/Lafter 25 minutes of homogenization. Typically, the mixture ishomogenized for an additional 5 minutes. Furthermore, the thermallability and sensitivity to oxidation of the sulfites necessitateaccurate temperature control and a nitrogen or other inert gasenvironment in the manufacturing process.

This procedure may be modified to prepare other compositions of thepresent invention by substituting other water immiscible solvents forthe soybean oil, other surfactants for the egg yolk phospholipid, otheracids or bases to adjust the pH instead of sodium hydroxide, and/orother tonicity modifiers for the glycerin. The procedure may also bemodified to prepare other drugs in a preserved oil-in-water emulsion orthose for parenteral nutrition.

3. Microbiological Activity

The growth retarding capability of 1% propofol injectable emulsioncontaining sodium metabisulfite, sodium bisulfite, sodium sulfite,potassium metabisulfite, or potassium sulfite were evaluated usingmembrane filtration technique and broth cultures. Approximately 50-200colony forming units (CFU) per mL of four standard organisms recommendedby United States Pharmacopeia (USP) for preservative efficacy tests wereinoculated in each formulation. These four organisms are identified asStaphylococcus aureus (ATCC 6538), Escherichia coli (ATCC 8739),Pseudomonas aeruginosa (ATCC 9027), and Candida albicans (ATCC 10231).In addition to these organisms, S. epidermidis (ATCC 12228) and S.aureus (coagulase negative, ATCC 27734) were also tested.

The antimicrobial activity of propofol containing sodium metabisulfitewas compared with propofol containing 0.005% disodiumethylenediaminetetraacetic acid (Diprivan EDTA, trademark Zeneca), and acontrol propofol formulation lacking preservative. After the inoculationof the test organisms, test formulations were incubated at 30-35° C. Theviable count of the test organism was determined immediately followingthe inoculation and after 24 h of incubation at 30-35° C. Each datum forthe metabisulfite composition is the average of eight determinationsperformed on two fresh 20-mL vials, two 1-month stability 20-mL vials,two fresh 100-mL vials, and two 1-month stability 100-mL vials. TheDiprivan samples were from four fresh 50-mL vials. Unpreserved propofolsamples contained the same ingredients, except they contained nopreservatives. The preservative was considered effective if themicrobial growth was suppressed, or allowed for a no-more-than 10-foldincrease in growth as compared to the zero-hour viable count (count ofthe organism immediately following inoculation) of each of the testorganisms.

The following Tables 4-12 compare the antimicrobial effectiveness ofsodium metabisulfite and other sulfite formulations with that ofDiprivan and unpreserved propofol. These results indicate that sodiummetabisulfite and the other sulfite compounds are competent to preventthe significant growth of microorganisms for at least 24 h afteradventitious, extrinsic contamination.

TABLE 4 Comparison of microbial growth retarding activity of variousformulations against S. aureus (ATCC 6538). Viable count of survivorsDecrease in Formulation log₁₀ CFU/mL ± SD survivors (number of samples)0 h 24 h log₁₀ CFU/mL Propofol 2.08 ± 0.28 ND 2.08 metabisulfite (8)Diprivan EDTA (4) 2.37 ± 0.26 1.55 ± 0.58 0.82 Unpreserved Propofol 2.05.5 NA (2) NA: Not applicable. ND: No viable organisms detected in 1-mLaliquot. SD: standard deviation.

TABLE 5 Comparison of microbial growth retarding activity of variousformulations against S. epidermidis (ATCC 12228). Viable count ofsurvivors Decrease in Formulation log₁₀ CFU/mL ± SD survivors (number ofsamples) 0 h 24 h log₁₀ CFU/mL Propofol 2.27 ± 0.04 ND 2.27metabisulfite (8) Diprivan EDTA (4) 2.20 ± 0.34 1.05 ± 0.35 1.15Unpreserved Propofol 2.4 4.55 ± 0.07 NA NA: Not applicable. ND: Noviable organisms detected in 1-mL aliquot. SD: standard deviation.

TABLE 6 Comparison of microbial growth retarding activity against E.coli (ATCC 8739) or various formulations. Viable count of survivorsDecrease in Formulation log₁₀ CFU/mL ± SD survivors (number of samples)0 h 24 h log₁₀ CFU/mL Propofol 2.26 ± 0.05 2.28 ± 0.13 NA metabisulfite(8) Diprivan EDTA (4) 2.37 ± 0.09 0.275 ± 0.55  2.095 UnpreservedPropofol 2.3 7.25 ± 0.07 NA NA: Not applicable. ND: No viable organismsdetected in 1-mL aliquot. SD: standard deviation.

TABLE 7 Comparison of microbial growth retarding activity of variousformulations against P. aeruginosa (ATCC 8739). Viable count ofsurvivors Decrease in Formulation log₁₀ CFU/mL ± SD survivors (number ofsamples) 0 h 24 h log₁₀ CFU/mL Propofol 1.97 ± 0.26 ND 1.97metabisulfite (8) Diprivan EDTA (4) 1.97 ± 0.17 2.50 ± 0.47 NAUnpreserved Propofol 2.35 ± 0.07 6.8 NA NA: Not applicable. ND: Noviable organisms detected in 1-mL aliquot. SD: standard deviation.

TABLE 8 Comparison of microbial growth retarding activity against S.aureus (coagulase negative, ATCC 27734) of various formulations. Viablecount of survivors Decrease in Formulation log₁₀ CFU/mL ± SD survivors(number of samples) 0 h 24 h log₁₀ CFU/mL Propofol 2.28 ± 0.22 ND 2.28metabisulfite (8) Diprivan EDTA (4) 2.87 ± 0.05 2.07 ± 0.28 0.80Unpreserved Propofol 3.15 ± 0.07 7.05 ± 0.07 NA NA: Not applicable. ND:No viable organisms detected in 1-mL aliquot. SD: standard deviation.

TABLE 9 Comparison of microbial growth retarding activity against C.albicans (ATCC 10231) of various formulations. Viable count of survivorsDecrease in Formulation log₁₀ CFU/mL ± SD survivors (number of samples)0 h 24 h log₁₀ CFU/mL Propofol 2.42 ± 0.08 3.13 ± 0.22 NA metabisulfite(8) Diprivan EDTA (4) 2.30 ± 0.08 3.20 ± 0.28 NA Unpreserved Propofol2.3 5.10 ± 0.14 NA NA: Not applicable. ND: No viable organisms detectedin 1-mL aliquot. SD: standard deviation.

TABLE 10 Comparison of microbial growth retarding activity against E.coli (ATCC 8739) of various sulfite- containing propofol injectableemulsion formulations (0.025%). Viable count of survivors Decrease inlog₁₀ CFU/mL ± SD survivors Formulations 0 h 24 h log₁₀ CFU/mL Propofolinjectable emulsion with: Sodium bisulfite 2.3 1.7 0.6 2.3 2.2 0.1Sodium sulfite 2.3 1.3 1.0 2.3 1.4 0.9 Potassium 2.3 2.0 0.3metabisulfite 2.3 2.0 0.3 Potassium sulfite¹ 2.3 4.8 NA 2.3 4.7 NADiprivan (50 ml) 2.5 0.5 2.0 2.5 0.3 2.2 Propofol without 2.3 7.3 NAantimicrobial agent NA: Not applicable. ND: No viable organisms detectedin 1-mL aliquot. SD: standard deviation. ¹Formulation contained 0.03mg/mL or 0.003% potassium sulfite. The inventors believe that a higherlevel of potassium sulfite will improve the inhibition of microbialgrowth.

TABLE 11 Comparison of microbial growth retarding activity against C.albicans (ATCC 10231) of various sulfite-containing propofol injectableemulsion formulations (0.025%). Viable count of survivors Decrease inlog₁₀ CFU/mL ± SD survivors Formulations 0 h 24 h log₁₀ CFU/mL Propofolinjectable emulsion with: Sodium bisulfite 2.3 1.6 0.7 2.3 1.4 0.9Sodium sulfite 2.3 ND 2.3 2.3 ND 2.3 Potassium 2.2 1.8 0.4 metabisulfite2.3 2.0 0.3 Potassium sulfite² 2.5 4.5 NA 2.4 4.5 NA Diprivan (50 ml)2.5 3.4 NA 2.5 3.4 NA Propofol without 2.3 5.1 NA antimicrobial agentNA: Not applicable. ND: No viable organisms detected in 1-mL aliquot.SD: standard deviation. ²Formulation contained 0.03 mg/mL or 0.003%potassium sulfite. The inventors believe that a higher level ofpotassium sulfite will improve the inhibition of microbial growth.

TABLE 12 Comparison of microbial growth retarding activity against P.aeruginosa (ATCC 9027) of various sulfite-containing propofol injectableemulsion formulations (0.025%). Viable count of survivors Decrease inlog₁₀ CFU/mL ± SD survivors Formulations 0 h 24 h log₁₀ CFU/mL Propofolinjectable emulsion with: Sodium bisulfite 1.8 ND 1.8 1.8 ND 1.8 Sodiumsulfite 1.5 ND 1.5 1.4 ND 1.4 Potassium 2.4 ND 2.4 metabisulfite 2.4 ND2.4 Potassium sulfite³ 2.3 ND 2.3 2.3 ND 2.3 Diprivan (50 ml) 2.5 3.4 NA2.4 3.3 NA Propofol without 2.4 6.8 NA antimicrobial agent NA: Notapplicable. ND: No viable organisms detected in 1-mL aliquot. SD:standard deviation. ³Formulation contained 0.03 mg/mL or 0.003%potassium sulfite. In spite of this low concentration, the potassiumsulfite was still effective in inhibiting growth of P. aeruginosa.

The present invention provides a sterile pharmaceutical preparation ofpropofol that comprises an amount of sulfite sufficient to significantlyprevent the growth, or prevent no more than 10-fold increase in growthof each of S. aureus (ATCC 6538), E. coli (ATCC 8739), P. aeruginosa(ATCC 9027), and C. albicans (ATCC 10231) S. epidermidis (ATCC 12228)and S. aureus (coagulase negative, ATCC 27734). Preferably, the sulfiteis sodium metabisulfite, sodium bisulfite, sodium sulfite, potassiummetabisulfite, or potassium sulfite. Furthermore, in the event ofimproper aseptic handling of the finished product leading to anaccidental extrinsic contamination, the present formulation willsuppress, minimize, or limit the chance of microbial growth for at least24 h.

EXAMPLES

Preferred compositions are as follows: 1% propofol emulsion forinjection:

a. about 1% propofol;

b. about 10% weight soybean oil;

C. about 2.25% weight glycerin;

d. about 1.2% weight egg-yolk phospholipid;

e. about 0.025% weight sulfite;

f. sodium hydroxide;

g. water to 100%. 2% propofol emulsion for injection:

a. about 2% propofol;

b. about 10% weight soybean oil;

c. about 2.25% weight glycerin;

d. about 1.2% weight egg-yolk phospholipid;

e. about 0.025% weight sulfite;

f. sodium hydroxide;

g. water to 100%.

Preferably, these formulations have pH of approximately 4.5-6.4.

The above examples of compositions, and methods of manufacturing sameare exemplary and the invention is not limited solely to those examples.

We claim:
 1. A sterile, pharmaceutical composition for parenteraladministration which comprises an oil-in-water emulsion in whichpropofol is dissolved in a water-immiscible solvent, is emulsified withwater, and is stabilized by means of a surfactant, and which furthercomprises an amount of sulfite sufficient to prevent a no more than10-fold increase in the growth of each of Staphylococcus aureus (ATCC6538), Escherichia coli (ATCC 8739), Pseudomonas aeruginosa (ATCC 9027),and Candida albicans (ATCC 1023 1) for at least 24 h as measured by atest wherein a washed suspension of each organism is added to a separatealiquot of said composition at approximately 50 colony-forming units permL and incubated at a temperature in the range 30-35° C. and are testedfor viable counts of said organisms after 24 h, and wherein said amountof sulfite is in the range of about 0.0075% to about 0.66% by weight ofsaid composition.
 2. The sterile, pharmaceutical composition accordingto claim 1 wherein the sulfite is selected from the group consisting ofsodium metabisulfite, sodium sulfite, sodium bisulfite, potassiummetabisulfite, and potassium sulfite.
 3. The sterile, pharmaceuticalcomposition according to claim 2 wherein said sulfite is sodiummetabisulfite.
 4. The sterile, pharmaceutical composition according toclaim 1 which comprises from about 10% to about 20% weight of awater-immiscible solvent.
 5. The sterile, pharmaceutical compositionaccording to claim 1 wherein the pH is between about 4.5 to about 6.4.6. The sterile, pharmaceutical composition according to claim 1 whichcomprises from about 1% to about 2% weight propofol.
 7. A sterile,pharmaceutical composition for parenteral administration which comprisesan oil-in-water emulsion in which propofol is dissolved in awater-immiscible solvent, is emulsified with water, and is stabilized bymeans of a surfactant, and which further comprises an amount of sulfitewherein the amount of sulfite is in the range of about 0.0075% to about0.66% weight, wherein the pH of said composition is between about 4.5 toabout 6.4.
 8. The sterile, pharmaceutical composition according to claim7 which comprises from about 10% to about 20% weight of awater-immiscible solvent.
 9. The sterile, pharmaceutical compositionaccording to claim 7 which comprises from about 1% to about 2% weightpropofol.
 10. The sterile, pharmaceutical composition of claim 7 whereinthe sulfite is selected from the group consisting of sodiummetabisulfite, sodium sulfite, sodium bisulfite, potassiummetabisulfite, and potassium sulfite.
 11. The pharmaceutical compositionof claim 10 wherein said sulfite is sodium metabisulfite.
 12. A sterile,pharmaceutical composition for parenteral administration which comprisesan oil-in-water emulsion in which propofol is emulsified with water, andis stabilized by means of a surfactant, and which further comprises anamount of sulfite sufficient to prevent a no more than 10-fold increasein the growth of each of Staphylococcus aureus (ATCC 6538), Escherichiacoli (ATCC 8739), Pseudomonas aeruginosa (ATCC 9027), and Candidaalbicans (ATCC 10231) for at least 24 h as measured by a test wherein awashed suspension of each organism is added to a separate aliquot ofsaid composition at approximately 50 colony-forming units per mL andincubated at a temperature in the range 30-35° C. and are tested forviable counts of said organisms after 24 h, and wherein said amount ofsulfite is in the range of about 0.0075% to about 0.66% weight of saidcomposition.
 13. A sterile, total-parenteral-nutrition composition forparenteral administration which comprises an oil-in-water emulsion inwhich a lipid is emulsified with water and which further comprises anamount of sulfite sufficient to prevent a no more than 10-fold increasein the growth of each of Staphylococcus aureus (ATCC 6538), Escherichiacoli (ATCC 8739), Pseudomonas aeruginosa (ATCC 9027), and Candidaalbicans (ATCC 1023 1) for at least 24 h as measured by a test wherein awashed suspension of each organism is added to a separate aliquot ofsaid composition at approximately 50 colony-forming units per mL andincubated at a temperature in the range 30-35° C. and are tested forviable counts of said organisms after 24 h, wherein said amount ofsulfite is in the range of about 0.0075% to about 0.66% weight of saidcomposition.
 14. A sterile, pharmaceutical composition for parenteraladministration which comprises an oil-in-water emulsion in which alipophilic pharmaceutical or therapeutic agent is dissolved in awater-immiscible solvent, is emulsified with water, and is stabilized bymeans of a surfactant, and which further comprises an amount of sulfitesufficient to prevent a no more than 10-fold increase in the growth ofeach of Staphylococcus aureus (ATCC 6538), Escherichia coli (ATCC 8739),Pseudomonas aeruginosa (ATCC 9027), and Candida albicans (ATCC 1023 1)for at least 24 h as measured by a test wherein a washed suspension ofeach organism is added to a separate aliquot of said composition atapproximately 50 colony-forming units per mL and incubated at atemperature in the range 30-35° C. and are tested for viable counts ofsaid organisms after 24 h, and wherein said amount of sulfite is in therange of about 0.0075% to about 0.66% weight of said composition.
 15. Asterile, pharmaceutical composition for parenteral administration whichcomprises an oil-in-water emulsion in which a lipophilic pharmaceuticalor therapeutic agent is emulsified with water and is stabilized by meansof a surfactant, and which further comprises an amount of sulfitesufficient to prevent a no more than 10-fold increase in the growth ofeach of Staphylococcus aureus (ATCC 6538), Escherichia coli (ATCC 8739),Pseudomonas aeruginosa (ATCC 9027), and Candida albicans (ATCC 1023 1)for at least 24 h as measured by a test wherein a washed suspension ofeach organism is added to a separate aliquot of said composition atapproximately 50 colony-forming units per mL and incubated at atemperature in the range 30-35° C. and are tested for viable counts ofsaid organisms after 24 h, and wherein said amount of sulfite is in therange of about 0.0075% to about 0.66% weight said composition.
 16. Amethod for inducing anesthesia comprising parenteral administration of acomposition which comprises an oil-in-water emulsion in which propofolis dissolved in a water-immiscible solvent, is emulsified with water,and is stabilized by means of a surfactant, and which further comprisesan amount of sulfite sufficient to prevent a no more than 10-foldincrease in the growth of each of Staphylococcus aureus (ATCC 6538),Escherichia coli (ATCC 8739), Pseudomonas aeruginosa (ATCC 9027), andCandida albicans (ATCC 1023 1) for at least 24 h as measured by a testwherein a washed suspension of each organism is added to a separatealiquot of said composition at approximately 50 colony-forming units permL and incubated at a temperature in the range 30-35° C. and are testedfor viable counts of said organisms after 24 h, and wherein said amountof sulfite is in the range of about 0.0075% to about 0.66% weight ofsaid composition.
 17. The method for inducing anesthesia according toclaim 16 wherein the method of administration is by continuous infusion.18. A method of maintaining anesthesia comprising parenteraladministration of a composition which comprises an oil-in-water emulsionin which propofol is dissolved in a water-immiscible solvent, isemulsified with water, and is stabilized by means of a surfactant, andwhich further comprises an amount of sulfite sufficient to prevent a nomore than 10-fold increase in the growth of each of Staphylococcusaureus (ATCC 6538), Escherichia coli (ATCC 8739), Pseudomonas aeruginosa(ATCC 9027), and Candida albicans (ATCC 1023 1) for at least 24 h asmeasured by a test wherein a washed suspension of each organism is addedto a separate aliquot of said composition at approximately 50colony-forming units per mL and incubated at a temperature in the range30-35° C. and are tested for viable counts of said organisms after 24 h,and wherein said amount of sulfite is in the range of about 0.0075% toabout 0.66% weight of said composition.
 19. The method of maintaininganesthesia according to claim 18 wherein the method of administration isby continuous infusion.
 20. A method of sedation comprising parenteraladministration of a composition which comprises an oil-in-water emulsionin which propofol is dissolved in a water-immiscible solvent, isemulsified with water, and is stabilized by means of a surfactant, andwhich further comprises an amount of sulfite sufficient to prevent a nomore than 10-fold increase in the growth of each of Staphylococcusaureus (ATCC 6538), Escherichia coli (ATCC 8739), Pseudomonas aeruginosa(ATCC 9027), and Candida albicans (ATCC 1023 1) for at least 24 h asmeasured by a test wherein a washed suspension of each organism is addedto a separate aliquot of said composition at approximately 50colony-forming units per mL and incubated at a temperature in the range30-35° C. and are tested for viable counts of said organisms after 24 h,and wherein said amount of sulfite is in the range of about 0.0075% toabout 0.66% weight of said composition.
 21. The method of sedationaccording to claim 20 wherein the method of administration is bycontinuous infusion.
 22. A sterile pharmaceutical composition accordingto any one of claim 12, 13, 14 or 15 wherein said sulfite is sodiummetabisulfite.
 23. A method according to any one of claim 17, 19 or 21wherein said sulfite is selected from the group consisting of sodiummetabisulfite, sodium sulfite, sodium bisulfite, potassium sulfite,potassium bisulfite, and potassium metabisulfite, and wherein the amountof said sulfite is in the range of about 0.0075% to about 0.66% weight.24. A method according to claim 23 wherein said sulfite is sodiummetabisulfite.
 25. A method according to claim 24 wherein saidcomposition comprises from about 10% to about 20% weight of awater-immiscible solvent.
 26. A method according to claim 24 wherein thepH of the emulsion is between about 4.5 to about 6.4.
 27. A methodaccording to claim 24 wherein said composition comprises from about 1%to about 2% weight propofol.